Factor VIII (FVIII) is a protein found in blood plasma, which acts as a cofactor in the cascade of reactions leading to blood coagulation. A deficiency in the amount of FVIII activity in the blood results in the clotting disorder known as hemophilia A, an inherited condition primarily affecting males. Hemophilia A is currently treated with therapeutic preparations of FVIII derived from human plasma or manufactured using recombinant DNA technology. Such preparations are administered either in response to a bleeding episode (on-demand therapy) or at frequent, regular intervals to prevent uncontrolled bleeding (prophylaxis).
FVIII is known to be relatively unstable in therapeutic preparations. In blood plasma, FVIII is usually complexed with another plasma protein, von Willebrand factor (vWF), which is present in plasma in a large molar excess to FVIII and is believed to protect FVIII from premature degradation. Another circulating plasma protein, albumin, may also play a role in stabilizing FVIII in vivo. Currently marketed FVIII preparations therefore primarily rely on the use of albumin and/or vWF to stabilize FVIII during the manufacturing process and during storage.
The albumin and vWF used in currently marketed FVIII preparations are derived from human blood plasma, however, and the use of such material has certain drawbacks. Because a large molar excess of albumin compared to FVIII is generally added in order to increase the stability of the FVIII in such preparations, it is difficult to characterize the FVIII protein itself in these preparations. The addition of human-derived albumin to FVIII is also perceived as being a disadvantage with respect to recombinantly-produced FVIII preparations. This is because, in the absence of such added albumin, the theoretical risk of transmitting a virus would be reduced in recombinantly-derived FVIII preparations.
Several attempts to formulate FVIII without albumin or vWF (or with relatively low levels of these excipients) have been described. For example, U.S. Pat. No. 5,565,427 (EP 508 194) to Freudenberg (assigned to Behringwerke) describes FVIII preparations which contain particular combinations of detergent and amino acids, specifically arginine and glycine, in addition to excipients such as sodium chloride and sucrose. The detergent, polysorbate 20 or polysorbate 80, is described as being present in amounts of between 0.001 to 0.5% (v/v), while arginine and glycine are present in amounts of between 0.01 to 1 mol/l. Sucrose is described as being present in amounts of between 0.1 and 10%. Example 2 of this patent alleges that solutions of (1) 0.75% sucrose, 0.4 M glycine, and 0.15M NaCl, and (2) 0.01 M sodium citrate, 0.08 M glycine, 0.016M lysine, 0.0025 M calcium chloride, and 0.4 M sodium chloride were not stable in solution over 16 hours, whereas solutions of (3) 1% sucrose, 0.14 M arginine, 0.1 M sodium chloride and (4) 1% sucrose, 0.4 M glycine, 0.14 M arginine, 0.1 M sodium chloride, and 0.05% TWEEN™. 80 (polysorbate 80) exhibited stability.
U.S. Pat. No. 5,763,401 (EP 818 204) to Nayer (assigned to Bayer) also describes a therapeutic FVIII formulation without albumin, comprising 15-60 mM sucrose, up to 50 mM NaCl, up to 5 mM calcium chloride, 65-400 mM glycine, and up to 50 mM histidine. The following specific formulations were identified as allegedly being stable: (1) 150 mM NaCl, 2.5 mM calcium chloride, and 165 mM mannitol; and (2) 1% sucrose, 30 mM sodium chloride, 2.5 mM calcium chloride, 20 mM histidine, and 290 mM glycine. A formulation containing higher amounts of sugar (10% maltose, 50 mM NaCl, 2.5 mM calcium chloride, and 5 mM histidine) was allegedly found to exhibit poor stability in the lyophilized state compared with formulation (2).
U.S. Pat. No. 5,733,873 (EP 627 924) to Osterberg (assigned to Pharmacia & Upjohn) discloses formulations which include between 0.01-1 mg/ml of a surfactant. This patent discloses formulations having the following ranges of excipients: polysorbate 20 or 80 in an amount of at least 0.01 mg/ml, preferably 0.02-1.0 mg/ml; at least 0.1 M NaCl; at least 0.5 mM calcium salt; and at least 1 mM histidine. More particularly, the following specific formulations are disclosed: (1) 14.7, 50, and 65 mM histidine, 0.31 and 0.6 M NaCl, 4 mM calcium chloride, 0.001, 0.02, and 0.025% polysorbate 80, with or without 0.1% PEG 4000 or 19.9 mM sucrose; and (2) 20 mg/ml mannitol, 2.67 mg/ml histidine, 18 mg/ml NaCl, 3.7 mM calcium chloride, and 0.23 mg/ml polysorbate 80.
Other attempts to use low or high concentrations of sodium chloride have also been described. U.S. Pat. No. 4,877,608 (EP 315 968) to Lee (assigned to Rhone-Poulenc Rorer) discloses formulations with relatively low concentrations of sodium chloride, namely formulations comprising 0.5 mM to 15 mM NaCl, 5 mM calcium chloride, 0.2 mM to 5 mM histidine, 0.01 to 10 mM lysine hydrochloride and up to 10% sugar. The “sugar” can be up to 10% maltose, 10% sucrose, or 5% mannitol.
U.S. Pat. No. 5,605,884 (EP 0 314 095) to Lee (assigned to Rhone-Poulenc Rorer) teaches the use of formulations with relatively high concentrations of sodium chloride. These formulations include 0.35 M to 1.2 M NaCl, 1.5 to 40 mM calcium chloride, 1 mM to 50 mM histidine, and up to 10% of a “sugar” such as mannitol, sucrose, or maltose. A formulation comprising 0.45 M NaCl, 2.3 mM calcium chloride, and 1.4 mM histidine is exemplified.
International Patent Application WO 96/22107 to Roser (assigned to Quadrant Holdings Cambridge Limited) describes formulations which include the sugar trehalose. These formulations comprise: (1) 0.1 M NaCl, 15 mM calcium chloride, 15 mM histidinetr, and 1.27 M (48%) ehalose; or (2) 0.011% calcium chloride, 0.12% histidine, 0.002% Tris, 0.002% TWEEN™. 80, 0.004% PEG 3350, 7.5% trehalose, and either 0.13% or 1.03% NaCl.
U.S. Pat. No. 5,328,694 (EP 511 234) to Schwinn (assigned to Octapharma AG) describes a formulation which includes 100 to 650 mM disaccharide and 100 mM-1.0 M amino acid. Specifically, the following formulations are disclosed: (1) 0.9 M sucrose, 0.25 M glycine, 0.25 M lysine, and 3 mM calcium chloride; and (2) 0.7 M sucrose, 0.5 M glycine, and 5 mM calcium chloride.
Other therapeutic FVIII formulations of the prior art generally include albumin and/or vWF for the purpose of stabilizing FVIII and are therefore not relevant to the present disclosure. Although there exists an extensive literature focused on formulation and process development issues with freeze dried products, studies of freeze dried FVIII are limited to a study of formulations based upon use of NaCl as a bulking agent
A major complication in formulation development is the presence of sodium chloride in the bulk drug substance, which is generally introduced by the purification process. As a result of the purification process, large and/or variable amounts of sodium chloride are present in the bulk drug substance solution. Uncrystallized sodium chloride lowers the collapse temperature and may leave the product incapable of being manufactured in anything resembling an elegant product. Further, the amorphous sodium chloride may compromise stability of the product. The present disclosure involves formulations for freeze drying wherein NaCl is removed or present in trace amounts, which are capable of keeping FVIII stably stored for extended periods of time.